Friedreich's ataxia: past, present and future

Friedreich's ataxia (FRDA) is an autosomal recessive inherited disorder characterized by progressive gait and limb ataxia, dysarthria, areflexia, loss of vibratory and position sense, and a progressive motor weakness of central origin. Additional features include hypertrophic cardiomyopathy and diabetes. Large GAA repeat expansions in the first intron of the FXN gene are the most common mutation underlying FRDA. Patients show severely reduced levels of a FXN-encoded mitochondrial protein called frataxin. Frataxin deficiency is associated with abnormalities of iron metabolism: decreased iron-sulfur cluster (ISC) biogenesis, accumulation of iron in mitochondria and depletion in the cytosol, enhanced cellular iron uptake. Some models have also shown reduced heme synthesis. Evidence for oxidative stress has been reported. Respiratory chain dysfunction aggravates oxidative stress by increasing leakage of electrons and the formation of superoxide. In vitro studies have demonstrated that Frataxin deficient cells not only generate more free radicals, but also show a reduced capacity to mobilize antioxidant defenses. The search for experimental drugs increasing the amount of frataxin is a very active and timely area of investigation. In cellular and in animal model systems, the replacement of frataxin function seems to alleviate the symptoms or even completely reverse the phenotype. Therefore, drugs increasing the amount of frataxin are attractive candidates for novel therapies. This review will discuss recent findings on FRDA pathogenesis, frataxin function, new treatments, as well as recent animal and cellular models. Controversial aspects are also discussed.     

Pathophysiogical and therapeutic progress in Friedreich ataxia

Friedreich ataxia (FRDA) is the most common hereditary autosomal recessive ataxia, but is also a multisystemic condition with frequent presence of cardiomyopathy or diabetes. It has been linked to expansion of a GAA-triplet repeat in the first intron of the FXN gene, leading to a reduced level of frataxin, a mitochondrial protein which, by controlling both iron entry and/or sulfide production, is essential to properly assemble and protect the Fe-S cluster during the initial stage of biogenesis. Several data emphasize the role of oxidative damage in FRDA, but better understanding of pathophysiological consequences of FXN mutations has led to develop animal models. Conditional knockout models recapitulate important features of the human disease but lack the genetic context, GAA repeat expansion-based knock-in and transgenic models carry a GAA repeat expansion but they only show a very mild phenotype. Cells derived from FRDA patients constitute the most relevant frataxin-deficient cell model as they carry the complete frataxin locus together with GAA repeat expansions and regulatory sequences. Induced pluripotent stem cell (iPSC)-derived neurons present a maturation delay and lower mitochondrial membrane potential, while cardiomyocytes exhibit progressive mitochondrial degeneration, with frequent dark mitochondria and proliferation/accumulation of normal mitochondria. Efforts in developing therapeutic strategies can be divided into three categories: iron chelators, antioxidants and/or stimulants of mitochondrial biogenesis, and frataxin level modifiers. A promising therapeutic strategy that is currently the subject of intense research is to directly target the heterochromatin state of the GAA repeat expansion with histone deacytelase inhibitors (HDACi) to restore frataxin levels.     

Normal serum iron and ferritin concentrations in patients with Friedreich's ataxia

Friedreich's ataxia (FRDA) is caused by point mutations or trinucleotide repeat expansions in both alleles of the gene encoding frataxin. Studies of frataxin homologues in lower eukaryotes suggest that mitochondrial iron accumulation may underlie the pathophysiology of FRDA. To evaluate the possible role of iron-chelation therapy for FRDA, we measured serum iron and ferritin concentration in 10 FRDA patients. The measurements were within normal limits, suggesting that iron-chelation therapy for FRDA may be problematic.     

Friedreich's ataxia with chorea and myoclonus caused by a compound heterozygosity for a novel deletion and the trinucleotide GAA expansion.

Friedreich's ataxia (FRDA) is the most common hereditary ataxia, affecting about 1 in 50,000 individuals. It is caused by mutations in the frataxin gene; 98% of cases have homozygous expansions of a GAA trinucleotide in intron 1 of the frataxin gene. The remaining 2% of patients are compound heterozygotes, who have a GAA repeat expansion in one allele and a point mutation in the other allele. FRDA patients with point mutation have been suggested to have atypical clinical features. We present a case of compound heterozygotes in a FRDA patient who has a deletion of one T in the start codon (ATG) of the frataxin gene and a GAA repeat expansion in the other allele. The patient presented with chorea and subsequently developed FRDA symptoms. The disease in this case is the result of both a failure of initiation of translation and the effect of the expansion. This novel mutation extends the range of point mutations seen in FRDA patients, and also broadens the spectrum of FRDA genotype associated with chorea.     

Iron metabolism in mice with partial frataxin deficiency

Friedreich ataxia (FRDA), the most common autosomal recessive inherited ataxic disorder, is the consequence of deficiency of the mitochondrial protein frataxin, typically caused by homozygous intronic GAA expansions in the corresponding gene. The yeast frataxin homologue (yfh1p) is required for cellular respiration. Yfh1p appears to regulate mitochondrial iron homeostasis and protect from free radical toxicity. Complete loss of frataxin in knockout mice leads to early embryonic lethality, indicating an important role for frataxin during development. Heterozygous littermates with partial frataxin deficiency are apparently healthy and have no obvious phenotype. Here we evaluate iron metabolism and sensitivity to dietary and parenteral iron loading in heterozygote frataxin knockout mice (Fx(+/-)). Iron concentrations in the liver, heart, pancreas and spleen, and cellular iron distribution patterns were compared between wild type and Fx(+/-) mice. Response to parenteral iron challenge was not different between Fx(+/-) mice and wild type littermates, while sporadic iron deposits were observed in the hearts of dietary iron-loaded Fx(+/-) mice. Finally, we evaluated the effect of partial frataxin deficiency on susceptibility to cardiac damage in the mouse model of hereditary hemochromatosis (HH), the Hfe knockout mice. HH, an iron overload disease, is one of the most frequent genetic diseases in populations of European origin. By breeding Hfe(-/-) with Fx(+/-) mice, we obtained compound mutant mice lacking both Hfe and one frataxin allele. Sparse iron deposits in areas of mild to moderate cardiac fibrosis were found in the majority of these mice. However, they did not develop any neurological symptoms. Our studies indicate an association between frataxin deficiency, iron deposits and cardiac fibrosis, but no obvious association between iron accumulation and neurodegeneration similar to FRDA could be detected in our model. In addition, these results suggest that frataxin mutations may have a modifier role in HH, that predisposes to cardiomyopathy.     

Friedreich ataxia: The clinical picture

Friedreich ataxia (FRDA) is a rare autosomal recessive hereditary disorder that affects approximately 1 in 50,000 Caucasians. It is caused by hyperexpansion of GAA repeats in the first intron of the frataxin gene. Initial symptoms of FRDA usually appear around the beginning of the second decade of life. In addition to neuropathological disabilities such as ataxia, sensory loss, and muscle weakness, common signs are scoliosis, foot deformity, and hypertrophic cardiomyopathy. Approximately 10 % of patients with FRDA develop diabetes. The neuronopathy in the dorsal root ganglia, accompanied by the loss of peripheral sensory nerve fibres and the degeneration of posterior columns of the spinal cord, is a hallmark of the disease and is responsible for the typical combination of signs and symptoms specific to FRDA. Variation in neurophysiological abnormalities is correlated with the size of the GAA repeat expansion and likely accounts for individual variation in the progression of FRDA. Despite a range of disease severity, most patients will lose their ability to walk, stand, or sit without support within 10 to 15 years of disease onset. In addition to a review of the clinicopathological features of FRDA, a discussion of recent advances in our understanding of the underlying molecular mechanisms is provided.     

Quantitative profiling and identification of differentially expressed plasma proteins in friedreich's ataxia

Friedreich's ataxia (FRDA) is an autosomal recessive ataxia, characterized by progressive gait ataxia, limb ataxia, dysarthria, and areflexia associated with diabetes and hypertrophic cardiomyopathy. The primary cause of FRDA is the presence of expanded DNA triplet (GAA) repeats in the first intron of the fxn gene on chromosome 9q13. The expanded DNA repeats in fxn inhibit expression of the protein frataxin, which leads to neuronal degeneration. The aim of the study was to identify differentially expressed plasma proteins in FRDA patients for their diagnostic/prognostic applications. Clinically suspected FRDA patients (n = 42) were assessed on the International Co‐Operative Ataxia Rating Scale (ICARS), and genetic confirmation was performed by analyzing (GAA) repeats via PCR. Eighteen patients were confirmed to be homozygous for FRDA, with ICARS scores of 40 ± 8. Plasma proteomics of homozygous FRDA patients and age‐ and gender‐matched healthy controls was done using two‐dimensional difference in‐gel electrophoresis and LC‐MS/MS. Quantitative proteomic analysis (fold change ≥1.5; P < 0.05) revealed 13 differentially expressed protein spots. These proteins were found to be associated with neuropathy (α1‐antitrypsin), ataxia (apolipoprotein A‐I), oxidative stress (albumin), altered lipid metabolism (apolipoprotein C‐II, C‐III), etc. Further investigations of these differentially expressed proteins can aid in identifying prognostic/diagnostic markers for FRDA. © 2013 Wiley Periodicals, Inc.     

Gene Expression Profile in Peripheral Blood Cells of Friedreich Ataxia Patients

Friedreich ataxia (FRDA) is the most common autosomal recessive ataxia characterized by a combination of neurological involvement, cardiomyopathy, and skeletal and glucose metabolism disturbances. FRDA is caused by mutations in FXN gene that results in reduction of mRNA and protein levels of frataxin. Previous microarray and real-time quantitative PCR (qPCR) studies showed that the downregulation of FXN is associated with a complex gene expression profile. However, these studies showed a wide variability in the subset of genes with altered expression among tissues and models. Genes differentially expressed in peripheral blood cells (PBC) could potentially help in the understanding of FRDA pathophysiology and also function as reliable disease biomarkers obtained from an easily accessible tissue, which could have implications in clinical practice. This study aimed to validate by qPCR the expression of 26 genes, revealed as differentially expressed by other studies, using peripheral blood cells (PBC) of 11 FRDA patients compared to 11 healthy controls. We found a robust downregulation of FXN, but no statistically significant differences were found between FRDA and controls for the remaining genes. Except for FXN, our study did not find a differential gene expression profile in PBC of FRDA patients and a reliable gene expression profile biomarker in a clinical relevant and noninvasive tissue remains unclear.     

Progress in the treatment of Friedreich ataxia

Friedreich ataxia (FRDA) is a progressive neurological disorder affecting approximately 1 in 29,000 individuals of European descent. At present, there is no approved pharmacological treatment for this condition however research into treatment of FRDA has advanced considerably over the last two decades since the genetic cause was identified. Current proposed treatment strategies include decreasing oxidative stress, increasing cellular frataxin, improving mitochondrial function as well as modulating frataxin controlled metabolic pathways. Genetic and cell based therapies also hold great promise. Finally, physical therapies are being explored as a means of maximising function in those affected by FRDA.     

Effects of Erythropoietin on Frataxin Levels and Mitochondrial Function in Friedreich Ataxia �C a Dose�CResponse Trial

Friedreich ataxia (FRDA) is an autosomal recessive inherited neurodegenerative disorder leading to reduced expression of the mitochondrial protein frataxin. Previous studies showed frataxin upregulation in FRDA following treatment with recombinant human erythropoietin (rhuEPO). Dose-response interactions between frataxin and rhuEPO have not been studied until to date. We administered escalating rhuEPO single doses (5,000, 10,000 and 30,000?IU) in monthly intervals to five adult FRDA patients. Measurements of frataxin, serum erythropoietin levels, iron metabolism and mitochondrial function were carried out. Clinical outcome was assessed using the "Scale for the assessment and rating of ataxia". We found maximal erythropoietin serum concentrations 24?h after rhuEPO application which is comparable to healthy subjects. Frataxin levels increased significantly over 3?months, while ataxia rating did not reveal clinical improvement. All FRDA patients had considerable ferritin decrease. NADH/NAD ratio, an indicator of mitochondrial function, increased following rhuEPO treatment. In addition to frataxin upregulation in response to continuous low-dose rhuEPO application shown in previous studies, our results indicate for a long-lasting frataxin increase after single high-dose rhuEPO administration. To detect frataxin-derived neuroprotective effects resulting in clinically relevant improvement, well-designed studies with extended time frame are required.     

Frataxin point mutations in two patients with Friedreich's ataxia and unusual clinical features

Two patients with a progressive ataxia are presented with clinical features consistent with classic Friedreich's ataxia (FRDA), but also with features unusual for FRDA. Analysis of DNA showed that each patient is heterozygous for the expanded GAA repeat of FRDA, but carries a base change on his other frataxin allele. For one patient a non-conservative arginine to cysteine amino acid change is predicted at amino acid 165 whereas the other mutation is found at the junction of exon one and intron one. Muscle biopsy showed an absence of frataxin immunoreactivity in the patient harbouring the intronic mutation, confirming the pathological nature of the base change. These mutations extend the range of point mutations seen in FRDA, and agree with recent reports suggesting phenotypic variation in patients with FRDA harbouring point mutations in conjunction with an expanded GAA repeat.     

Variations of frataxin protein levels in normal individuals

Friedreich’s ataxia (FRDA) is the most common of the inherited ataxias and is associated with GAA trinucleotide repeat expansions within the first intron of the frataxin (FXN) gene. There are expanded FXN alleles from 66 to 1,700 GAA·TTC repeats in FRDA patients and correlations between number of GAA repeats and frataxin protein levels are assumed. Here, we present for the first time frataxin protein levels as well as analysis of GAA triplet repeats in the FXN gene in a population of 50 healthy Austrian people. Frataxin protein levels were measured in lymphocytes from blood samples by ELISA and GAA repeats were analyzed by capillary electrophoresis. Rather unexpectedly, we found a high variation of frataxin protein levels among the individuals. In addition, there was no correlation between frataxin levels, GAA repeats, age and sex in this group. However, these findings are of great importance for better characterization of the disease.     

Mapping of the second Friedreich's ataxia (FRDA2) locus to chromosome 9p23-p11: evidence for further locus heterogeneity

Friedreich's ataxia (FRDA), the most-common form of autosomal recessive ataxia, is inherited in most cases by a large expansion of a GAA triplet repeat in the first intron of the frataxin (X25) gene. Genetic heterogeneity in FRDA has been previously reported in typical FRDA families that do not link to the FRDA locus on chromosome 9q13. We report localization of a second FRDA locus (FRDA2) to chromosome 9p23-9p11, and we provide evidence for further genetic heterogeneity of the disease, in a family with the classic FRDA phenotype.     

Exploring mental status in Friedreich's ataxia: a combined neuropsychological, behavioral and neuroimaging study

Despite much evidence of cognitive and affective disorders in Friedreich's ataxia (FRDA), the nature of mental status in FRDA has received little systematic attention. It has been proposed that the cerebellum may interfere indirectly with cognition through the cerebello-cortical loops, whereas the role of pathological changes in different areas of the central nervous system is still undetermined. In the present study, 13 patients with molecularly determined FRDA and a group of matched controls were evaluated by a comprehensive battery of neuropsychological tests and the Minnesota Multiphasic Personality Inventory. A repetitive task of simple visual-reaction times was used to investigate implicit learning in all subjects. Pathological changes in cortical areas were explored comparing cerebral activations of patients and controls during finger movements (functional MRI). The intelligence profile of FRDA patients is characterized by concrete thinking, poor capacity in concept formation and visuospatial reasoning. FRDA patients show reduced speed of information processing. The learning effect seen in controls was notably absent in patients with FRDA. The patients' personality is characterized by some pathological aspects and reduced defensiveness. Patterns of cortical activation during finger movements are heterogeneous in patients compared to controls. Cognitive impairment, mood disorders and motor deficits in FRDA patients may be the result of the cumulative damage caused by frataxin deficiency not only in the cerebellum and spinal cord but also in other brain areas.     

Multicellular models of Friedreich ataxia

Patients with Friedreich ataxia (FRDA) have severely reduced levels of the mitochondrial protein frataxin, which results from a large GAA triplet-repeat expansion within the frataxin gene (FXN). High evolutionary conservation of frataxin across species has enabled the development of disease models of FRDA in various unicellular and multicellular organisms. Mouse models include classical knockout models, in which the Fxn gene is constitutively inactivated, and knock-in models, in which a GAA repeat mutation or the conditional allele is inserted into the genome. Recently, “humanised” GAA repeat expansion mouse models were obtained by combining the constitutive knockout with the transgenic expression of a yeast artificial chromosome carrying the human FRDA locus. In lower organisms such as Caenorhabditis elegans and Drosophila, straight-forward and conditional RNA interference technology has provided an easy way to knock down frataxin expression. Conditional mouse models have been used for pre-clinical trials of potential therapeutic agents, including idebenone, MnTBAP (a superoxide dismutase mimetic), and iron chelators. Various models of FRDA have shown that different, even opposite, phenotypes can be observed, depending on the level of frataxin expression. Additional studies with animal models will be essential for an enhanced understanding of the disease pathophysiology and for the development of better therapies.     

Iron–sulfur cluster synthesis,iron homeostasis and oxidative stress in Friedreich ataxia

Friedreich ataxia (FRDA) is an autosomal recessive, multi-systemic degenerative disease that results from reduced synthesis of the mitochondrial protein frataxin. Frataxin has been intensely studied since its deficiency was linked to FRDA in 1996. The defining properties of frataxin – (i) the ability to bind iron, (ii) the ability to interact with, and donate iron to, other iron-binding proteins, and (iii) the ability to oligomerize, store iron and control iron redox chemistry – have been extensively characterized with different frataxin orthologs and their interacting protein partners. This very large body of biochemical and structural data [reviewed in (Bencze et al., 2006)] supports equally extensive biological evidence that frataxin is critical for mitochondrial iron metabolism and overall cellular iron homeostasis and antioxidant protection [reviewed in (Wilson, 2006)]. However, the precise biological role of frataxin remains a matter of debate. Here, we review seminal and recent data that strongly link frataxin to the synthesis of iron–sulfur cluster cofactors (ISC), as well as controversial data that nevertheless link frataxin to additional iron-related processes. Finally, we discuss how defects in ISC synthesis could be a major (although likely not unique) contributor to the pathophysiology of FRDA via (i) loss of ISC-dependent enzymes, (ii) mitochondrial and cellular iron dysregulation, and (iii) enhanced iron-mediated oxidative stress. This article is part of a Special Issue entitled ‘Mitochondrial function and dysfunction in neurodegeneration’.     

A novel deletion–insertion mutation identified in exon 3 of <Emphasis Type="Italic">FXN</Emphasis> in two siblings with a severe Friedreich ataxia phenotype

Friedreich ataxia (FRDA) is an autosomal recessive neurodegenerative disease most commonly caused by a GAA trinucleotide repeat expansion in the first intron of FXN, which reduces expression of the mitochondrial protein frataxin. Approximately 98% of individuals with FRDA are homozygous for GAA expansions, with the remaining 2% compound heterozygotes for a GAA expansion and a point mutation within FXN. Two siblings with early onset of symptoms experienced rapid loss of ambulation by 8 and 10 years. Diagnostic testing for FRDA demonstrated one GAA repeat expansion of 1010 repeats and one non-expanded allele. Sequencing all five exons of FXN identified a novel deletion-insertion mutation in exon 3 (c.371_376del6ins15), which results in a modified frataxin protein sequence at amino acid positions 124–127. Specifically, the amino acid sequence changes from DVSF to VHLEDT, increasing frataxin from 211 residues to 214. Using the known structure of human frataxin, a theoretical 3D model of the mutant protein was developed. In the event that the modified protein is expressed and stable, it is predicted that the acidic interface of frataxin, known to be involved in iron binding and interactions with the iron–sulphur cluster assembly factor IscU, would be impaired.